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Fig. 4 | Molecular Cancer

Fig. 4

From: DNA-methylation-mediated activating of lncRNA SNHG12 promotes temozolomide resistance in glioblastoma

Fig. 4

DNA methylation and SP1 are involved in the activation of SNHG12. a Schematic representation of the CpG islands and bisulfite sequencing; Magenta words, CG sites for bisulfite sequencing; Red region, input sequence; Blue region, CpG islands; BSP1 F1 and R1, BSP2 F1 and R1, bisulfite forward primer and reverse primer. b Bisulfite genomic sequencing was performed to examine methylation status of CpG island 1 at the promoter region of SNHG12 in NHA, Pri GBM, Rec GBM, N3S, N3T3rd cells. c MSP analysis was performed to examine methylation status of CpG island 1 at the promoter region of SNHG12 in normal brain tissues, primary GBM tissues and recurrent GBM tissues. d qRT-PCR analysis detecting the SNHG12 levels in Pri GBM and N3S cells after the treatment with 5-azacytidine for 72 h and 144 h. e The luciferase reporter plasmids carrying SNHG12 promoter region were co-transfected into HEK293T cells with five transcription factor plasmids, respectively. Relative luciferase activity in HEK293T cells were determined. f The SNHG12 levels were detected in Rec GBM and Pri GBM cells either stably expressing SP1 or with SP1 depleted. g The correlation between SP1 and SNHG12 in GBM tissues was analyzed. h Predicted SP1-binding sites in the promoter region of SNHG12. i ChIP-PCR assay of the enrichment of SP1 on the SNHG12 promoter region. j ChIP analysis for the detection of SP1 binding to the promoter region of SNHG12. k DNMT profiles using western blotting. l MSP analysis of CpG island 1 after DNMT1 overexpression. m qRT-PCR analysis of SNHG12 expression after DNMT1 overexpression or knockdown. n ChIP-PCR assay of the enrichment of SP1 on the SNHG12 promoter region after DNMT1 overexpression or knockdown. Data are presented as the mean ± SEM from three independent experiments. Significant results were presented as NS non-significant, *P<0.05, **P<0.01, ***P<0.001

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